Proteinuria is an invariable finding in patients with most types of chronic kidney disease and it is one of the few modifiable risk factors for long-term prognosis and progression to end stage renal failure. Regardless of the original etiology, the pathologic basis for glomerular proteinuria is podocyte dysfunction or injury. Evidence suggests that glycogen synthase kinase (GSK) 3, a multi-tasking kinase, plays an important role in mediating acute and chronic injuries in multiple solid organs including the kidney through regulating multiple pathogenic cellular events, such as mitochondria permeability transition (MPT), proinflammatory NFB activation, and cytoskeleton disorganization. Inhibition of GSK3 prevents kidney injury and represents a novel renoprotective strategy. The studies proposed here will decipher the putative role of GSK3 in podocyte dysfunction and test the novel hypothesis that blockade of GSK3 protects podocyte from injury, improves proteinuria and slow progression of glomerulosclerosis. Aim 1 will examine the role of GSK3 in adriamycin induced podocyte injury in cultured podocytes. GSK3 activity in podocytes will be specifically manipulated by RNA interference or ectopic expression of either inactive or non-inhibitable mutant GSK3. The regulatory effect of GSK3 on adriamycin induced podocyte injuries will be assessed, including MPT and the ensuing podocyte death, proinflammatory NFB activation and de novo expression of the costimulatory molecule B7-1, an NFB target gene, as well as podocyte shape changes and the underlying cytockeleton disorganization;Aim 2 will determine the effect of doxycycline inducible podocyte specific GSK3 knockout on adriamycin induced podocytopathy and proteinuria in adult mice. These studies are essential to conclusively elucidate the role of GSK3 in podocyte injury in vivo because selective GSK3 inhibitors may have nonspecific effects and podocyte specific blockade of GSK3 is impossible pharmacologically. Pathogenic mechanisms identified in Aim1 by which GSK3 promotes podocyte injury will be validated in vivo in the knockout mice. Aim 3 will test the preventive and rescue effects of TDZD-8, a novel non-ATP competitive small molecule inhibitor of GSK3 on adriamycin induced nephropathy. The efficacy of TDZD-8 will be compared with low dose lithium, a safe and effective FDA approved drug that possesses potent GSK3 inhibitory actions, already exists for decades and could be used for clinical trials years before kidney specific GSK3 blockade is possible. The effects of TDZD-8 or lithium on adriamycin induced podocyte injury and related mechanisms will be delineated. Collectively, these studies should allow rapid progress to clinical trials of existing drugs with GSK3 inhibitory activities to improve podocyte injury, induce proteinuria remission, and slow progression of glomerulosclerosis in man. PUBLIC HEALTH RELEVANCE: Proteinuria is an invariable finding in patients with most types of chronic kidney disease and is the final common pathway driving progression to glomerulosclerosis and end stage renal failure. The pathologic basis for glomerular proteinuria is podocyte dysfunction and injury. Existing strategies to improve proteinuria, including glucocorticoids and cytotoxic drugs, have severe side effects and limited utility. The studies proposed here will provide a comprehensive picture of a new cellular signaling molecule important in the regulation of podocyte injury and investigate an entirely novel therapeutic approach to attenuate proteinuria based on blocking this molecule. These studies should allow rapid progress to clinical trials of existing drugs with blocking activities to improve podocyte injury, induce proteinuria remission, and slow progression of glomerulosclerosis in man.